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. 1983 Apr;3(4):580–586. doi: 10.1128/mcb.3.4.580

Fusion of the Saccharomyces cerevisiae leu2 gene to an Escherichia coli beta-galactosidase gene.

A E Martinez-Arias, M J Casadaban
PMCID: PMC368573  PMID: 6406836

Abstract

The promoter and translation initiation region of the Saccharomyces cerevisiae leu2 gene was fused to the Escherichia coli beta-galactosidase gene. This fusion located the control region of the leu gene and orientated its direction of expression. When the fusion was placed into yeast cells, beta-galactosidase was expressed under the same regulatory pattern as the original leu2 gene product: its synthesis was repressed in the presence of leucine and threonine. Sensitive chromogenic substrates for beta-galactosidase were used to detect expression in isolated colonies growing on agar medium. Mutant yeast cells with increased beta-galactosidase activity were identified by the color of the colonies they formed. One class of mutants obtained appeared to affect ars1 plasmid maintenance, and another class appeared to affect beta-galactoside uptake.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Andreadis A., Hsu Y. P., Kohlhaw G. B., Schimmel P. Nucleotide sequence of yeast LEU2 shows 5'-noncoding region has sequences cognate to leucine. Cell. 1982 Dec;31(2 Pt 1):319–325. doi: 10.1016/0092-8674(82)90125-8. [DOI] [PubMed] [Google Scholar]
  2. Beggs J. D. Transformation of yeast by a replicating hybrid plasmid. Nature. 1978 Sep 14;275(5676):104–109. doi: 10.1038/275104a0. [DOI] [PubMed] [Google Scholar]
  3. Bennetzen J. L., Hall B. D. The primary structure of the Saccharomyces cerevisiae gene for alcohol dehydrogenase. J Biol Chem. 1982 Mar 25;257(6):3018–3025. [PubMed] [Google Scholar]
  4. Botstein D., Falco S. C., Stewart S. E., Brennan M., Scherer S., Stinchcomb D. T., Struhl K., Davis R. W. Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments. Gene. 1979 Dec;8(1):17–24. doi: 10.1016/0378-1119(79)90004-0. [DOI] [PubMed] [Google Scholar]
  5. Casadaban M. J., Chou J., Cohen S. N. In vitro gene fusions that join an enzymatically active beta-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translational initiation signals. J Bacteriol. 1980 Aug;143(2):971–980. doi: 10.1128/jb.143.2.971-980.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Casadaban M. J., Cohen S. N. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. J Mol Biol. 1980 Apr;138(2):179–207. doi: 10.1016/0022-2836(80)90283-1. [DOI] [PubMed] [Google Scholar]
  7. Casadaban M. J., Cohen S. N. Lactose genes fused to exogenous promoters in one step using a Mu-lac bacteriophage: in vivo probe for transcriptional control sequences. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4530–4533. doi: 10.1073/pnas.76.9.4530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Casadaban M. J., Martinez-Arias A., Shapira S. K., Chou J. Beta-galactosidase gene fusions for analyzing gene expression in escherichia coli and yeast. Methods Enzymol. 1983;100:293–308. doi: 10.1016/0076-6879(83)00063-4. [DOI] [PubMed] [Google Scholar]
  9. Casadaban M. J. Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. J Mol Biol. 1976 Jul 5;104(3):541–555. doi: 10.1016/0022-2836(76)90119-4. [DOI] [PubMed] [Google Scholar]
  10. Dickson R. C. Expression of a foreign eukaryotic gene in Saccharomyces cerevisiae: beta-galactosidase from Kluyveromyces lactis. Gene. 1980 Sep;10(4):347–356. doi: 10.1016/0378-1119(80)90155-9. [DOI] [PubMed] [Google Scholar]
  11. Franklin N. C. Genetic fusions for operon analysis. Annu Rev Genet. 1978;12:193–221. doi: 10.1146/annurev.ge.12.120178.001205. [DOI] [PubMed] [Google Scholar]
  12. Hinnen A., Hicks J. B., Fink G. R. Transformation of yeast. Proc Natl Acad Sci U S A. 1978 Apr;75(4):1929–1933. doi: 10.1073/pnas.75.4.1929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hsu Y. P., Kohlhaw G. B. Overproduction and control of the LEU2 gene product, beta-isopropylmalate dehydrogenase, in transformed yeast strains. J Biol Chem. 1982 Jan 10;257(1):39–41. [PubMed] [Google Scholar]
  14. Kohlhaw G. B., Hsu Y. P., Lemmon R. D., Petes T. D. Transposed LEU2 gene of Saccharomyces cerevisiae is regulated normally. J Bacteriol. 1980 Nov;144(2):852–855. doi: 10.1128/jb.144.2.852-855.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lindegren G., Hwang Y. L., Oshima Y., Lindegren C. C. Genetical mutants induced by ethyl methanesulfonate in Saccharomyces. Can J Genet Cytol. 1965 Sep;7(3):491–499. doi: 10.1139/g65-064. [DOI] [PubMed] [Google Scholar]
  16. Müller-Hill B., Kania J. Lac repressor can be fused to beta-galactosidase. Nature. 1974 Jun 7;249(457):561–563. doi: 10.1038/249561a0. [DOI] [PubMed] [Google Scholar]
  17. Ratzkin B., Carbon J. Functional expression of cloned yeast DNA in Escherichia coli. Proc Natl Acad Sci U S A. 1977 Feb;74(2):487–491. doi: 10.1073/pnas.74.2.487. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Rose M., Casadaban M. J., Botstein D. Yeast genes fused to beta-galactosidase in Escherichia coli can be expressed normally in yeast. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2460–2464. doi: 10.1073/pnas.78.4.2460. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Satyanarayana T., Umbarger H. E., Lindegren G. Biosynthesis of branched-chain amino acids in yeast: regulation of leucine biosynthesis in prototrophic and leucine auxotrophic strains. J Bacteriol. 1968 Dec;96(6):2018–2024. doi: 10.1128/jb.96.6.2018-2024.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Schümperli D., Howard B. H., Rosenberg M. Efficient expression of Escherichia coli galactokinase gene in mammalian cells. Proc Natl Acad Sci U S A. 1982 Jan;79(2):257–261. doi: 10.1073/pnas.79.2.257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Silverman S. J., Rose M., Botstein D., Fink G. R. Regulation of HIS4-lacZ fusions in Saccharomyces cerevisiae. Mol Cell Biol. 1982 Oct;2(10):1212–1219. doi: 10.1128/mcb.2.10.1212. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Zaret K. S., Sherman F. DNA sequence required for efficient transcription termination in yeast. Cell. 1982 Mar;28(3):563–573. doi: 10.1016/0092-8674(82)90211-2. [DOI] [PubMed] [Google Scholar]

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